Sys Rev Pharm 2020; 11(6): 420 424 A multifaceted review journal in the field of pharmacy E-ISSN 0976-2779 P-ISSN 0975-8453

Risk Assessment Metabolic Disorders by Prolonged Exposure to Low Doses of Benzene N.V. Sharapova1, S.I. Krasikov2, A.A. Petrova3, V.M. Boev4 1Candidate of biological Sciences, associate Professor of the Orenburg state medical University of the Ministry of Health of the Russian Federation, E-mail: [email protected] 2Doctor of medical Sciences, Professor of the Orenburg state medical University of the Ministry of Health of the Russian Federation, E-mail: [email protected] 3Assistant at the Orenburg state medical University of the Ministry of Health of the Russian Federation E-mail: [email protected] 4Doctor of medical Sciences, Professor of the Orenburg state medical University of the Ministry of Health of the Russian Federation, E-mail: [email protected]

Article History: Submitted: 12.04.2020 Revised: 16.05.2020 Accepted: 21.06.2020 ABSTRACT Benzene is one of the most common environmental pollutants. functions under the influence of benzene. In addition, there was a However, the main attention of researchers is drawn to its decrease in the level of calcium, sodium and potassium in the blood carcinogenic and hematotoxic effect. The metabolic effects of this serum, which may reflect the effect of the pollutant on the processes hydrocarbon have been studied to a lesser extent and are mainly of reabsorption of electrolytes in the renal tubules on the one hand, related to high effects. The associated with the effect on the as well as on the regulation of electrolyte . In addition, body of non-toxic doses of this pollutant are practically not studied, under the influence of low doses of benzene in animals developed which was the purpose of the present study. insulin resistance (IR), estimated by glucose tolerance test (GTT). The Experiments lasting 7 weeks were carried out on animals that possible role of oxidative stress in the development of the described received drinking water containing benzene at a of 10 disorders is discussed. µg/l, which is 1 maximum permissible concentration (MPC). Key words: benzene, , drinking water, maximum permissible It was shown that long-term consumption of water containing concentration, serum, total protein, uric acid, blood glucose, insulin benzene in concentration equal to 1 MPC in animals led to changes in resistance, enzymemia, dyslipoproteinemia biochemical parameters in blood serum, characterized by a decrease Correspondence: in the level of total protein and albumin, a moderate increase in the N. V. Sharapova activity of serum enzymes – alanine aminotransferase (ALAT), Candidate of Biological sciences, Associate Professor of the Orenburg aspartate aminotransferase (ASAT), alkaline phosphatase (ALP) and state medical university of the ministry of health of the Russian lactate dehydrogenase (LDH). Simultaneously, developed Federation hypercholesterolemia, triacylglycerolemia and dislipoproteinemia. In E-mail: [email protected] addition, there was a decrease in the concentration of urea and uric DOI: 10.31838/srp.2020.6.67 acid. The detected changes are considered as changes in liver @Advanced Scientific Research. All rights reserved

INTRODUCTION not been studied enough, and the results obtained are According to the International Program on Chemical Safety ambiguous, which is the reason for this work [14, 15, 16]. (IPCS), benzene is one of the ten most hazardous substances for [1] and is one of the most common MATERIALS AND METHODS environmental pollutants [2]. At the same time, the vast The work was performed on 24 male rats of the Wistar line. majority of studies assess the negative impact of benzene on The of the animals at the beginning of the experiment human health in terms of its hemotoxicity and was 170 grams. The animals were kept in a vivarium, with carcinogenicity [3, 4], as there is more and more information day-night cycles of 12/12 hours. We used granulated food about the data on its toxicity to various organs and systems from Procorm (Novosibirsk, Russia) for feeding, balanced in in literature[5], rather than the carcinogenic effects of nutrients, vitamins and microelements. The animals were benzene. In particular, the toxicity of benzene in relation to given bottled water extracted from local artesian springs the cardiovascular system [6], its hypertensive effect «Akva vita» (LLC "Zhivaya Planeta", Orenburg, Russia) for associated with a violation of the formation of nitrogen oxide drinking. The amount of feed and water was not limited. [7] is shown. Administration of benzene to animals had a All experimental animals were divided into two equal groups hepatotoxic effect, which appeared by hepatospecific of equal weight and size. Animals of the first group served as hyperfermentemia [8], as well as nephrotoxic effect control group. Benzene was added to rats of the second group accompanied by depression of carbohydrate metabolism to a concentration of 0.01 mg / liter in drinking water, which enzymes in the cortical and cerebral substance of the kidneys corresponded to 1 maximum permissible concentration [9]. (MPC). The duration of the experiment was 8 weeks. An important manifestation of benzene toxicity can be Half of the animals from each group were tested for glucose observed when it acts on the human and animal body insulin tolerance (TSH) at the 6th week of the experiment. For this, resistance [10, 11, 12], as well as violations of the expression after 12 hours of starvation, but maintaining access to water, of fatty acid metabolism enzymes [13]. each was injected per os with a pipette 20% glucose It should be noted that, as a rule, observations related to both at the rate of 2 grams of dry matter per kg of mass. carcinogenic and non-carcinogenic effects of benzene relate Immediately before administration, and then every 30 to high doses of the toxicant, while the data available in minutes after administration of glucose, blood was collected literature indicate the need to study the effect on the body of from the tail vein of the animals, in which the glucose this aromatic hydrocarbon at low exposures. However, the concentration was determined using a glucose meter and test negative effects of benzene on the body in small doses have strips "Accu-Chek performa" (Roshe, Switzerland).

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All animals were removed from the experiment in Statistical processing was performed by using the "Microsoft compliance with ethical standards after 8 weeks. The blood Excel" package, the reliability of differences was evaluated by was collected in test tubes containing a coagulation activator the Student's test. of the company "Sarshted" (Germany) after decapitation and by centrifugation at 3000 rpm, the serum was separated from RESULTS the shaped elements. The results of the experiments are presented in table 1. Biochemical studies of blood serum were carried out at the From the data provided it follows that, in rats consumed "Cobas-6000" station of Roche (Switzerland) and included water containing benzene at a concentration of 1 MPC, there the determination of the level of total protein, albumin, was a decrease in the concentration of protein in the blood creatinine, uric acid, activity of aspartic and alanine serum by 7%, and the albumin content was lower by 10% than transaminases ALAT and ASAT, alkaline phosphatase in the control group. The urea content in the experimental (ALP), lactate dehydrogenase (LDH), the concentration of and control groups was within the normal range, but at the total cholesterol (TC) and its fractions high-density same time, it was lower in animals receiving benzene by 10%. lipoprotein cholesterol (HDL), lipoproteins very high low The level of serum creatinine in the groups did not differ, and and low density (VLDL and LDL), triacylglycerol (TAG), the uric acid in rats treated with benzene was 1.6 times lower ferritin, sodium, potassium, calcium, ferrum. than in intact ones.

Table 1: Biochemical parameters of blood serum in rats with the use of water containing benzene (M ± m) Intact Benzene Indicators, units. P

n=12 n=12

Total protein, g/l 63,0±1,2 58,9±0,56 P1-2= 0,0055

Albumin, g/l 35,8±0,86 32,4±0,33 P1-2= 0,0014

Urea, mmol/l 5,9±0,13 5,4±0,21 P1-2= 0,055

Creatinine, mmol/l 35,3±1,61 35,8±1,57 P1-2=0,826

Uric acid, mmol/l 131,5±10,1 82,1±8,1 P1-2= 0,001

ASAT, units/l 132,5±6,9 171,0±12,0 P1-2= 0,011

ALAT, unit/l 51,7±2,5 59,03±3,1 P1-2= 0,080

ALP, units/l 150,6±3,0 166,0±6,0 P1-2= 0,032

LDH, units/l 547,0±21,0 669,0±37,0 P1-2= 0,009

TC, mol/l 1,61±0,1 1,5±0,06 P1-2= 0,356

HDL cholesterol, mol/l 1,54±0,06 1,31±0,03 P1-2= 0,003

LDL cholesterol, mol/l 0,28±0,034 0,25±0,04 P1-2= 0,574

VLDL cholesterol, mol/l 0,4±0,025 0,49±0,035 P1-2= 0,049

AI, standard unit 0,045±0,006 0,13±0,04 P1-2= 0,048

TG, mol/l 1,03±0,08 1,2±0,05 P1-2= 0,086

NEFA, mol/l 0,68±0,02 0,83±0,04 P1-2=0,003

Sodium, mmol/l 146,5±0,69 141,4±1,72 P1-2= 0,012

Potassium, mmol/l 6,09±0,25 5,26±0,17 P1-2= 0, 012 Calcium, mmol/l 2,5±0,04 2,3±0,03 P1-2= 0,001

Ferrum, µmol/l 23,5±1,29 19,12±0,39 P1-2= 0,004

Ferritin, µg/l 125±9,34 140±5,68 P1-2= 0,184

Further, the table shows that the intake of small doses of As a result, the Atherogenic Index (AI) in rats of the benzene into the bodies of animals led to an increase in the experimental series was 2.7 times higher than in intact activity of serum enzymes, while the activity of ASAT in the animals. experimental group of rats was 21% higher than in the The level of TAG and free FA in benzene-treated rats was 16 control group, ALAT by 14%, ALP by 10.6%, and LDH by and 23%, respectively, higher than in the control group. 22%. Further, the table shows that the content of the main serum Long-term intake of small doses of benzene did not lead to cations-Na+, K+ and Ca2+ - in animals receiving benzene was changes in the level of total cholesterol in the blood serum, lower by 6, 15 and 9%, respectively, compared to rats but had a significant effect on its distribution by fractions. consuming clean water. The table shows that the systemic exposure of HDL Figure 1 shows that fasting glucose levels in intact and cholesterol in experimental animals was lower than in the experimental animals did not differ. control by 18%.

421 Systematic Review Pharmacy Vol 11, Issue 6, 2020 N. V. Sharapova et al / Risk Assessment Metabolic Disorders by Prolonged Exposure to Low Doses of Benzene

Fig. 1: The concentration of glucose in the blood of rats during the glucose tolerance test

When conducting a glucose tolerance test, changes in its atherogenic index, as well as an increase in the concentration concentration in the blood of intact animals were of TAG. The lower than in the control, the concentration of characterized by a rise to 30 minutes after oral electrolytes in blood serum observed in rats treated with low administration, and then a decrease almost to the initial doses of benzene, can reflect as a violation of the reabsorption values after 2 hours (Fig. 1). processes in the renal tubules, this is more true for sodium The curve showing changes in glucose concentration in and potassium ions, and on the other hand, a decrease in animals receiving benzene was characterized, firstly, by absorption in enterocytes which is possible for calcium ions. higher values per 30 minute after oral administration, and Finally, long-term consumption of water containing low secondly, by a slower decrease in its level in subsequent of benzene led to the development of insulin periods. At the same time, by the 180th minute of the test, resistance, assessed by the glucose tolerance test. the serum glucose values in the experimental group were 1.25 In other words, long-term intake of benzene in low doses led mmol/l higher than the control group. to changes in both biochemical parameters and metabolic regulation processes. DISCUSSION The obtained data are in accordance with studies that reflect Thus, the results obtained indicate that prolonged intake of the effects of low effects of benzene on the human body [3, benzene in small doses with drinking water leads to 16, 17, 18]. noticeable changes in the metabolic status of animals. The most likely reason for the effects of low doses of benzene Firstly, these changes were characterized by a decrease of the described, in our opinion, is the peculiarities of its level of total protein occurring due to the albumin fraction. metabolism when entering the body. Thus, according to At the same time, the benzene-treated rats had lower levels of modern concepts, this aromatic hydrocarbon undergoes urea and uric acid than in the control group. These listed biotransformation in the endoplasmic reticulum of the liver, symptoms may indicate a slight decrease in the metabolic with the participation of the isoform of cytochrome P450- function of the liver. First of all, it is subject to the processes 2E1 [19, 20]. of albumin synthesis, as well as the formation of urea and The transformations that occur with its participation are catabolism of purine nucleotides. characterized by the formation of reactive and It should be noted that these changes, which are not toxic metabolites [19] and lead to the activation of free radical significant in their severity, but have a significant character, oxidation of membrane , both in the liver and in other occur against the background of moderate organs. In other words, the metabolism of benzene entering hyperfermentemia, estimated by the activity of ASAT, ALAT, the body is accompanied by the development of oxidative ALP and LDH, which may be evidence of increased stress, which is known to cause damage to the membrane permeability of hepatocyte membranes. structures of cells, as well as proteins and nucleic acids, which Changes in liver function can probably be associated with the is an important basis for mutagenesis. It should be noted that observed dyslipoproteinemia, which is manifested by a the nature of transformations does not depend on the decrease in the proportion of the antiatherogenic fraction of amount of incoming benzene. cholesterol represented by HDL and an increase in the

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ABOUT THE AUTHORS Natalia Sharapova (Orenburg, Russia) candidate of biological Sciences, associate Professor of the Orenburg state medical University of the Ministry of Health of the Russian Federation, (e-mail: [email protected]; tel.: 8 (3532) 600505). Sergey Krasikov (Orenburg, Russia) - doctor of medical Sciences, Professor of the Orenburg state medical University of the Ministry of Health of the Russian Federation, (e-mail: [email protected]; tel.: 8 (3532) 600505). Alyona Petrova (Orenburg, Russia) - assistant at the Orenburg state medical University of the Ministry of Health of the Russian Federation, (e-mail: [email protected]; tel.: 8 (3532) 600505). Viktor Boev (Orenburg, Russia) - doctor of medical Sciences, Professor of the Orenburg state medical University of the Ministry of Health of the Russian Federation, (e-mail: [email protected]; tel.: 8 (3532) 600505). Correspondent mailing address: Russia, 460000, Orenburg, st. Sovetskaya, 6.

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